Lipstatin is of considerable importance as key intermediate for the preparation of tetrahydrolipstatin (THL, Orlistat), which is useful in the prophylaxis and treatment of diseases associated with obesity.
Lipstatin, a fermentative process for its production, a process for its isolation from microorganisms and a process for its hydrogenation to tetrahydrolipstatin are known and described for example in U.S. Pat. No. 4,598,089.
Lipstatin is depicted as follows: ##STR1##
A process for the preparation of crude lipstatin has also been described in European Patent Application No. 96106598. This process comprises aerobically cultivating a microorganism of the order of actinomycetes, e.g. Streptomyces toxytricini, which produces lipstatin, in an aqueous medium which is substantially free of fats and oils, and which contains suitable carbon and nitrogen sources and inorganic salts, until the initial growth phase is substantially finished and sufficient cell mass has been produced. Then, linoleic acid together with an antioxidant, and optionally together with caprylic acid, and N-formyl-L-leucine or preferably L-leucine is added to the broth. After completion of the fermentation, the fermentation broth is extracted. The produced crude lipstatin can be further enriched and purified, e.g. by chromatographic methods, described in U.S. Pat. No. 4,598,089.
Multiple step chromatographic protocols or liquid-liquid extractions to enrich lipstatin in combination with multiple step chromatography to obtain pure lipstatin characterizes prior-art methods for the purification of crude lipstatin. These methods are typically used for the isolation of fermentation metabolites on a laboratory scale. However, these methods are generally not suitable for an economic large-scale process.
Attempts to purify crude lipstatin by distillation failed. Lipstatin is stable for several hours at 60.degree. C., but because of its low vapor pressure (7.times.10.sup.-7 mbar) vacuum distillation (&lt;2 mbar) at this temperature is not feasible. At higher temperature lipstatin is degraded by elimination of carbon dioxide.
Crude lipstatin can also be purified by crystallization of lipstatin at temperatures below -20.degree. C. However expensive technical equipment for low temperature crystallization is required and yield of this crystallization strongly depends on the quality of crude lipstatin. Especially with low quality of crude lipstatin, lipstatin is obtained in low yield via crystallization.